University of Khartoum

MOLECULAR CHARACTERIZATION AND DRUG RESISTANCE PATTERNS OF MYCOBACTERIUM TUBERCULOSIS ISOLATES FROM KHARTOUM STATE

MOLECULAR CHARACTERIZATION AND DRUG RESISTANCE PATTERNS OF MYCOBACTERIUM TUBERCULOSIS ISOLATES FROM KHARTOUM STATE

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Title: MOLECULAR CHARACTERIZATION AND DRUG RESISTANCE PATTERNS OF MYCOBACTERIUM TUBERCULOSIS ISOLATES FROM KHARTOUM STATE
Author: Mukhtar, Moawia M.; ELHASSAN, ALI MOHAMMED
Abstract: The present study was an attempt to study the epidemiology of pulmonary tuberculosis, characterization and identification of the causative mycobacteria and determination of their susceptibility to rifampin as a marker for drug resistance in the Khartoum state, Sudan. One hundred and fifty patients were consented and selected for the study. All sputum samples examined showed the characteristic AFB appearance of serpentine cords. All smear positive samples showed colonies of growth characteristic for Mycobacterium tuberculosis and all of them showed the presence of AFB on indirect smearing. Demographic data of consented patients were obtained from 147 patients out of 150 patients. All age groups were found to be affected by tuberculosis with varying frequencies. Five patients (3.4%) from group one (5-10), 32 (21.8%) from group two (11-20), 43 (29.3%) from group three (21-30), 30 (21.8%) from group four (31-40), 19 (12.9%) from group five (41-50), 8 (5.4%) from group six (51-60), 6 (4.1%) from group seven (61-70), and 2 patients in group eight (1.4%). 98 (66.7%) of the enrolled patients were males while 49 (33.3%) were females. The disease was found to be prevalent nearly in all provinces and localities of Khartoum state. 54 (36.7%) patients were found to be resident in the different localities of Khartoum, 56 (38.1%), and 27 (18.4%) were found to live in Khartoum North or Omdurman respectively, while 10 (6.8%)patients were considered as non Khartoum residents. 77 (52.4%) patients were singles (59 males, 18 females), 68 (46.3%) were married (35 males, 33 females), while one (0.7%) was scored for each of the separated and divorced categories. 39 (26.5%) were students, 30 (20.4%) were labors, 34 (23.1%) were housewives, 31 (21.1%) were business persons, 5 (3.4%) were employees (2 officers, 2 teachers, and 1 policeman), 5 (3.4%) were farmers, and 3 (2.1%) were homeless. 45 Sudanese tribes were recorded in this study. Patients from Denka, Gaalin and Nuba tribes were found to be significantly more than those from other tribes. The cough was the most common complaint (143, 97.3%), while other symptoms percentages were as follows, fever (121, 82.3%), loss of weight (96, 65.3%), shortness of breath (86, 58.5%), and haemoptysis (13, 8.8%). On duplex PCR, 129 out of 135 clinical isolates investigated by molecular biology techniques showed 235 bp DNA amplicons, indicating that they were M. tuberculosis complex strains while six isolates showed no DNA amplicons at neither 235 bp nor 136 bp. On PCR restriction enzyme analysis of rpoB gene (351 bp) the conserved rpoB gene present in all mycobacteria was amplified in 134 of our clinical isolates examined so far in this investigation with the exception of only one isolate which showed no amplified DNA bands. On nested PCR based sequence analysis amplifications of the rpoB gene of the clinical isolates resulted in the amplification of the target gene in all the examined isolates (133), at205 bp and 157 bp respectively, confirming that all of them were M. tuberculosis species. The G+C% content of the obtained nucleotide sequences was determined. A mean of 68.56% of G+C% content was determined. On analysis of the detected rpoB gene mutations, genetic alterations within the 81 bp hot spot region of rpoB gene known to be associated with rifampicin resistance were detected in eleven (11/133, 8.27%) of the studied isolates, of which the mutation Ser531 → Leu was the more common one (6, 54.55%), while mutations Leu511 → Pro, Asp516 → Gly, Asp516 → Val, Leu521 → Leu, and His526 → Tyr occurred once with the rate of 9.09% for each
URI: http://hdl.handle.net/123456789/6337
Date: 2004


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